Project 7: Regulation and integration in bacterial cells (Laub) (#34-37) We elucidated the design principles of regulatory modules in bacterial cells, with a focus on the signal transduction systems that process information. Histidine kinases play a major role in regulating bacterial physiology. We developed and used system-level tools to map the connections between kinases and substrates in Caulobacter crescentus. Our finding that histidine kinases exhibit a large kinetic preference in vitro for their in vivo cognate substrates led to new techniques for rapidly mapping signal transduction pathways in bacteria. These allowed us to show that the Caulobacter cell cycle is driven by an integrated genetic circuit that uses regulated transcription, proteolysis, and phosphorylation to produce oscillations in activity of the master cell cycle regulator CtrA15. We also identified the first bona fide cell cycle checkpoint in Caulobacter, demonstrating that this key principle of regulation in eukaryotes is also used in prokaryotes. Finally, we used computation to predict which residues account for the specificity of kinase-substrate interactions, and made mutations that switched the specificity of one protein kinase (EnvZ) to that of several others. Michael Laub, the PI, was a Bauer Fellow and is now Assistant Professor of Biology at MIT. New project 7: Life-history constraints on developmental modules in plants (Queitsch) (#38) We asked how genetic variation, maternal effects and chaperone levels determine developmental trajectories and fitness of Arabidopsis thaliana. Plants are not motile and must adapt to the conditions they experience. Adaptation depends on their genetic constitution, non-Mendelian effects transmitted from their mothers, and chaperone levels. We have investigated three sources of variation in traits that control life history (flowering time, seed yield, etc.): cryptic genetic variation that is exposed by environmental stress, maternal effects, and allelic differences between different geographic populations. We previously showed that altering HspQO levels reveals cryptic genetic variation, have now mapped the loci for these traits and find that one of them identifies a sugardependent cyclin. We discovered that differences in the environment plants experience have profound effects on their progeny and that these differences are transmitted through the ovule but not through pollen. These maternal effects depend on light sensing by phytochromes and show complex interactions with the effects of reducing Hsp90 levels. Finally, we examined the variation between different geographic isolates; they show substantial variation, and much of the variation responds (either increasing or decreasing) to reducing Hsp90 activity24. This work has revealed the complex regulation of ecologically important developmental traits and the importance of taking an integrated approach to understanding them. Christine Queitsch, the project leader, was a Bauer Fellow, joined the Center for Modular Biology in 2006 and is now Assistant Professor of Genome Sciences at U Washington.